Bottom Line:
In all, 139 transcripts related to various transcription regulation factors (TFs) were unsuccessfully activated in the iSCNT embryos.Incompatibilities between mitochondrial DNA (mtDNA) and nuclear DNA revealed that the TOMM (translocase of outer mitochondrial membrane)/TIMM (translocase of inner mitochondrial membrane) complex-associated genes in BBNT embryos had the highest expression levels, while the PBNT embryos exhibited much lower expression rates.Improper degradation of maternal transcripts, incomplete activation of TFs and abnormal expression of genes associated with mitochondrial function in PBNT embryos likely contributed to incomplete reprogramming of the donor cell nuclei and therefore led to the developmental failure of these cloned embryos.

Background: Interspecies somatic cell nuclear transfer (iSCNT) has been regarded as a potential alternative for rescuing highly endangered species and can be used as a model for studying nuclear-cytoplasmic interactions. However, iSCNT embryos often fail to produce viable offspring. The alterations in normal molecular mechanisms contributing to extremely poor development are for the most part unknown.

Results: Przewalski's gazelle-bovine iSCNT embryos (PBNT) were produced by transferring Przewalski's gazelle fibroblast nuclei into enucleated bovine oocytes. The percentages of PBNT embryos that developed to morula/blastocyst stages were extremely low even with the use of various treatments that included different SCNT protocols and treatment of embryos with small molecules. Transcriptional microarray analyses of the cloned embryos showed that the upregulation of reprogramming-associated genes in bovine-bovine SCNT (BBNT) embryos was significantly higher than those observed in PBNT embryos (1527:643). In all, 139 transcripts related to various transcription regulation factors (TFs) were unsuccessfully activated in the iSCNT embryos. Maternal degradation profiles showed that 1515 genes were uniquely downregulated in the BBNT embryos, while 343 genes were downregulated in the PBNT embryos. Incompatibilities between mitochondrial DNA (mtDNA) and nuclear DNA revealed that the TOMM (translocase of outer mitochondrial membrane)/TIMM (translocase of inner mitochondrial membrane) complex-associated genes in BBNT embryos had the highest expression levels, while the PBNT embryos exhibited much lower expression rates.

Conclusions: Improper degradation of maternal transcripts, incomplete activation of TFs and abnormal expression of genes associated with mitochondrial function in PBNT embryos likely contributed to incomplete reprogramming of the donor cell nuclei and therefore led to the developmental failure of these cloned embryos.

Mentions:
There were 25 of the transcription regulator-associated genes co-upregulated in both BBNT and PBNT embryos (Figure 4B). One hundred thirty-nine transcripts associated with transcriptional regulation were upregulated in BBNT embryos, whereas these genes had only low expression levels in PBNT embryos (Figure 4A; Additional file9: Table S6). These genes contribute to multiple biological functions such as general transcription factors, mediator complexes, nuclear receptor subfamily kinase, anchor proteins, RNA polymerases, and zinc finger proteins. As an example, the basic transcription factors for RNA polymerase (e.g., TBP, TFIIB, TAF1D, SP1 and TAF2) were all upregulated in BBNT embryos, but only TBP and TAF2 were upregulated in PBNT embryos (Additional file10: Figure S4). Unsupervised hierarchical clustering revealed that the expression pattern of these transcription factors in PBNT embryos were more similar to the donor somatic cells, which indicates that the key transcription regulatory pathway in PBNT embryos did not activate as in BBNT embryos (Figure 3).Figure 4

Mentions:
There were 25 of the transcription regulator-associated genes co-upregulated in both BBNT and PBNT embryos (Figure 4B). One hundred thirty-nine transcripts associated with transcriptional regulation were upregulated in BBNT embryos, whereas these genes had only low expression levels in PBNT embryos (Figure 4A; Additional file9: Table S6). These genes contribute to multiple biological functions such as general transcription factors, mediator complexes, nuclear receptor subfamily kinase, anchor proteins, RNA polymerases, and zinc finger proteins. As an example, the basic transcription factors for RNA polymerase (e.g., TBP, TFIIB, TAF1D, SP1 and TAF2) were all upregulated in BBNT embryos, but only TBP and TAF2 were upregulated in PBNT embryos (Additional file10: Figure S4). Unsupervised hierarchical clustering revealed that the expression pattern of these transcription factors in PBNT embryos were more similar to the donor somatic cells, which indicates that the key transcription regulatory pathway in PBNT embryos did not activate as in BBNT embryos (Figure 3).Figure 4

Bottom Line:
In all, 139 transcripts related to various transcription regulation factors (TFs) were unsuccessfully activated in the iSCNT embryos.Incompatibilities between mitochondrial DNA (mtDNA) and nuclear DNA revealed that the TOMM (translocase of outer mitochondrial membrane)/TIMM (translocase of inner mitochondrial membrane) complex-associated genes in BBNT embryos had the highest expression levels, while the PBNT embryos exhibited much lower expression rates.Improper degradation of maternal transcripts, incomplete activation of TFs and abnormal expression of genes associated with mitochondrial function in PBNT embryos likely contributed to incomplete reprogramming of the donor cell nuclei and therefore led to the developmental failure of these cloned embryos.

Background: Interspecies somatic cell nuclear transfer (iSCNT) has been regarded as a potential alternative for rescuing highly endangered species and can be used as a model for studying nuclear-cytoplasmic interactions. However, iSCNT embryos often fail to produce viable offspring. The alterations in normal molecular mechanisms contributing to extremely poor development are for the most part unknown.

Results: Przewalski's gazelle-bovine iSCNT embryos (PBNT) were produced by transferring Przewalski's gazelle fibroblast nuclei into enucleated bovine oocytes. The percentages of PBNT embryos that developed to morula/blastocyst stages were extremely low even with the use of various treatments that included different SCNT protocols and treatment of embryos with small molecules. Transcriptional microarray analyses of the cloned embryos showed that the upregulation of reprogramming-associated genes in bovine-bovine SCNT (BBNT) embryos was significantly higher than those observed in PBNT embryos (1527:643). In all, 139 transcripts related to various transcription regulation factors (TFs) were unsuccessfully activated in the iSCNT embryos. Maternal degradation profiles showed that 1515 genes were uniquely downregulated in the BBNT embryos, while 343 genes were downregulated in the PBNT embryos. Incompatibilities between mitochondrial DNA (mtDNA) and nuclear DNA revealed that the TOMM (translocase of outer mitochondrial membrane)/TIMM (translocase of inner mitochondrial membrane) complex-associated genes in BBNT embryos had the highest expression levels, while the PBNT embryos exhibited much lower expression rates.

Conclusions: Improper degradation of maternal transcripts, incomplete activation of TFs and abnormal expression of genes associated with mitochondrial function in PBNT embryos likely contributed to incomplete reprogramming of the donor cell nuclei and therefore led to the developmental failure of these cloned embryos.